Introduction

Over 800 million people, approximately 10.7% of the global population, currently endure chronic undernourishment and hunger, with the prevalence of this issue increasing steadily at a rate of approximately 10%1. Of particular concern is the high incidence of malnutrition among children, which accounts for 45% of child deaths, totaling nearly 14 million per year. Alarmingly, one-third of all global food production is wasted, with perishable produce playing a significant role in this problem, as up to 50% of fruits grown in the field are discarded annually2,3. Despite the existence of a natural protective barrier on the fruit surface, the perishability of fruits resulting from factors such as microbial growth, oxidation, dehydration, and aging manifests in limited shelf-life and rapid decay4. Hence, it is crucial to develop effective techniques to address these challenges and prolong the shelf life of produce to reduce food wastage.

To extend the shelf-life of perishable fruits, a variety of strategies, including genetic modification and harmful chemical preservatives (such as formaldehyde, copper sulfate, potassium sorbate, and nitrite etc.), have been explored. While these methods have shown success in prolonging fruit shelf-life, further advancements are needed to ensure biosafety and ecological sustainability5. Cold chain storage, another efficient method for extending shelf life, is expensive, energy-intensive, and contributes to high CO2 emissions. Functional coating technologies, which utilize natural substances such as proteins, polysaccharides, and lipids in composite formulations, offer a promising alternative6. These coatings create a semi-permeable film on the fruit surface, forming a protective barrier that regulates gas exchange and inhibits microbial infiltration7,8,9. To enhance the protective qualities of these coatings, antibacterial and antioxidant agents10,11,12,13, including essential oils14 and metal oxide nanoparticles13, are frequently incorporated. Despite persistent efforts, the fruit preservation coating primarily faces three key challenges: (1) Water-soluble preservation solutions struggle to evenly distribute on hydrophobic fruit cuticles, leading to protective vulnerabilities. (2) The diverse chemical composition and microstructure of fruit cuticle layers pose great difficulty to the universal adhesion of conventional protection layers on a variety of fruit surfaces. (3) Challenges in synergizing preservation components hinder the achievement of a synergistic effect greater than the sum of its parts.

Phase-transitioned proteins differ from native proteins as they form protein aggregates with highly organized cross-β-sheet structures instead of the typical α-helix conformations15. This transition predominantly occurs at gas-liquid or solid-liquid interfaces, resulting in the formation of two-dimensional protein nanofilms16,17. Notably, the two-dimensional protein film exhibits exceptional adhesion properties to a diverse range of substrates18,19, and has been used in diverse applications such as underwater adhesives20, enhancing fertilizer bioavailability21,22, and creating stain-resistant coatings for fabrics23. By carefully selecting proteins with inherent bactericidal properties24, phase-transitioned proteins have the potential to adhere, sterilize, and integrate other beneficial natural substances on the cuticle of perishable fruit. In this regard, a universal and biocompatible fruit preservation strategy is highly expected.

Herein, we have developed a sustainable phase-transitioned protein-based coating that provides versatile fresh-keeping functions for fruit preservation (Fig. 1). We introduced amyloid-like protein (ALP) to enhance the adhesion stability of the fresh-keeping coating by integrating lysozyme to leverage its antimicrobial properties within a phase transition system. Through computer-aided molecular simulation, we demonstrated that ALP forms a robust adhesive on the outermost epidermal wax layer. The addition of sodium alginate (SA) and sulfuric acid-treated cellulose nanocrystals (CNC) further enhances the coating’s film-forming ability and gas barrier properties, resulting in a synergistic fresh-keeping effect among all three components. The resulting edible ALP coating displays a notable inhibition of microbial growth, fruit respiration, and moisture loss, thereby significantly extending the shelf-life of non-climacteric and climacteric post-harvest fruits by 2 ~ 5-fold, highlighting its universal fruit preservation properties while preserving 60–98% of their nutrition, flavor, and firmness. These results surpass those achieved by chemical preservation and cold chain methods, providing a safer and more effective alternative. Notably, our coating reduces carbon emissions by 90% compared to standard refrigeration practices while extending shelf-life by 2.5 times without compromising nutrient retention. With a low treatment cost of $0.09/kg fruit, the ALP concept presents an effective and sustainable approach to addressing global food waste by preventing post-harvest fruit loss.

Fig. 1: Schematic diagram illustrating the preparation of ALP coatings and their role in fruit preservation.
figure 1

Phase-transitioned lysozyme, sodium alginate, and cellulose nanocrystals work together to enhance the ALP coating’s adhesion stability, mechanical strength, water and oxygen barrier properties, antibacterial performance, and antioxidant capabilities, thereby extending the shelf life of fruits. The ALP coating is washable, edible, cost-effective, reduces carbon emissions, and provides universal applicability in fruit preservation.

Full size image

Results

Design and preparation of the adhesive fresh-keeping ALP coating

The natural cuticular wax found on post-harvest fruits serves as the outermost protective layer of the fruit’s epidermal cells, shielding the fruit from microbial threats and physical harm. To enhance the preservation capabilities of this wax layer, lysozyme with intrinsic bacteria-killing properties, was chosen as an adhesive agent to boost the antimicrobial properties and mechanical resilience of fruit preservation coatings. Given that the cuticular wax typically exhibits an amorphous structure on a mesoscopic scale, as evidenced by scanning electron microscope (SEM) images (Supplementary Fig. 1), we initiated the process by constructing a model of the assembled cuticular wax using molecular dynamics (MD) simulations, which involved utilizing a hydrophobic blend of long-chain fatty acids, their derivatives, and triterpenoids to mimic the composition of the cuticular wax4 (Supplementary Fig. 2). The simulation results revealed the rapid anchoring of lysozyme onto the cuticular wax within 40 ps, with complete adhesion achieved at 25 ns (Fig. 2a and Supplementary Movie 1). The final snapshot depicted 33 lysozyme residues establishing interactions with the cuticular wax, resulting in a substantial contact area of 38.62 nm2 and a high binding energy of − 2832.69 kJ/mol (Fig. 2b). To systematically evaluate the interfacial adhesion mechanism, we investigated amyloid-like lysozyme (PDB entry: 8QV8) as an alternative to phase-transitioned lysozyme through molecular dynamics simulations with cuticular wax substrates. The simulations revealed that amyloid-like lysozyme achieves rapid surface spreading on cuticular wax within 2 ps, culminating in full interfacial adhesion by 1 ns (Supplementary Figs. 3, 4). Notably, while the amyloid variant spans residues 26-100 (versus the full 126 residues in native lysozyme), it exhibits enhanced molecular interactions with the wax surface. Quantitative analysis demonstrated a significantly expanded contact interface (48.50 nm²) and elevated binding energy (− 3713.02 kJ/mol) compared to the native protein (Fig. 2b). These metrics represent a 26% increase in interfacial contact area and a 31% enhancement in binding energy relative to native lysozyme. Extending our prior investigations of native lysozyme-wax interfacial dynamics, these studies indicate native lysozyme may first attach to fruit wax via random coil structures, then recruit more molecules to the wax interface, ultimately undergoing disulfide bond cleavage to form β-sheet-rich amyloid aggregates (Fig. 2a and Supplementary Fig. 5). This structural reorganization correlates with the observed adhesion strengthening, consistent with our previous findings on coil-mediated surface attachment and disulfide rupture-driven adhesion enhancement17. Remarkably, amyloid-like lysozyme achieves rapid interfacial consolidation across diverse material surfaces, forming chemically resistant and mechanically robust protein adlayers. This phenomenon suggests their potential as protective coatings for perishable commodities—the strong lysozyme-wax interfacial adhesion could mitigate chemical degradation and physical abrasion in postharvest fruits under extreme environmental conditions.

Fig. 2: Adhesion between fruits and ALP.
figure 2

a Molecular dynamics (MD) simulations illustrate the interfacial adhesion between fruit wax and lysozyme as well as its amyloid-like variant. b Binding energy and contact area of lysozyme and amyloid-like lysozyme on the wax surface after simulation. c CD spectra of native lysozyme, PTL, and ALP. d The peeling strength of ALP coating and native protein adsorption layer during the 180° peeling test. Data are mean ± S.D. n  =  3 independent samples per group. e Optical images depicting ALP agent and native lysozyme solution interactions on fruit surfaces. f Sequential optical images showing a sessile ALP droplet on various fruit peels at time points t = 0 s and 60 s. Source data are provided as a Source Data file.

Full size image

To prepare the fresh-keeping coating, lysozyme was then phase-transitioned by introducing cysteine, which served as a reducing agent and triggered the transformation of the protein chains from α-helix to β-sheet (Fig. 2c), and consequently, the phase-transitioned lysozyme (PTL) underwent amyloid-like aggregation to serve as a universal adhesive basement on the fruit surface18. Sodium alginate (SA), a natural polysaccharide extracted from brown seaweed, was then utilized as a substrate for the final coatings to enhance film-forming characteristics. By further combining with the exceptional gas barrier capabilities of cellulose nanocrystals (CNC), an amyloid-like protein (ALP) adhesive coating consisting of PTL, SA, and CNC was finally developed, and an optimized formula was then obtained through a three-factor, four-level orthogonal experiment (Supplementary Table 1). Analysis of the Fourier transform infrared (FTIR) spectra revealed a substantial increase in the β-sheet structure within the amide I band of the ALP coating, indicating the presence of amyloid-like aggregation of lysozyme in the ALP system (Supplementary Fig. 6). Such protein aggregation was further confirmed by thioflavin T (ThT) and 1-anilino-8-naphthalenesulfonic acid (ANS) assays (Supplementary Fig. 6). Moreover, high-resolution X-ray photoelectron spectroscopy (XPS) spectra of the C1s region unveiled various functional groups exposed on the ALP coating, including aliphatic carbon, amines, hydroxyl, amides, thiols, and carboxyl groups (Supplementary Fig. 7). These functional groups originated from the amyloid-like protein aggregation, CNCs, and SA, facilitating intermolecular hydrogen bonding within the coating, as demonstrated by the shift to high binding energies in the C1s, O1s, and N1s spectra (Supplementary Fig. 7). These functional groups and corresponding intermolecular interactions are responsible for the coating mechanical fortification and multiplex binding of the coating with the cuticular wax, so as to provide a robust fresh-keeping adhesive protection layer for various fruit cuticles.

By using a 180° peeling test, it was found that the adhesion levels of the ALP coating on hydrophobic substrates exhibited a 5-fold increase compared to the native lysozyme adsorption layer on a blank substrate (Fig. 2d and Supplementary Fig. 8). This result highlighted that the incorporation of PTL significantly enhanced the interfacial adhesion of the coating on various surfaces due to the amyloid-like structure-mediated interfacial adhesion15,16,17. The adhesive ability of the ALP coating on three hydrophobic fruits was further explored using dyed native lysozyme solution as the control (Fig. 2e). A uniform liquid film was observed on the fruit surfaces after soaking in the ALP fresh-keeping agents, contrasting with sporadic water droplets seen on fruits soaked in dyed native lysozyme solution. The uniform ALP liquid film formation can be attributed to the favorable contact angle on the fruit surface. For instance, the contact angle of the ALP agent on a cherry tomato was measured at 80°, decreasing to 40° within 60 s, while a similar phenomenon was also observed on other fruits, such as winter jujube, strawberry, kumquat, and banana (Fig. 2f). This result indicated the effective spreading ability of the ALP agents on perishable fruits. By further determining the surface tension and the work of adhesion (Wa) of the ALP agents (Supplementary Table 2), it is thereby evident that the ALP can seamlessly wet the cuticular wax of fruits and establish robust adhesion on a wide range of fruit surfaces.

Versatile fresh-keeping features of the ALP coating

The ideal fresh-keeping coating should possess multifunctional qualities, including rapid film formation, robust mechanical properties, appropriate adhesion strength, oxidation resistance, and antimicrobial activity. Initially, the ALP coating demonstrates shear-thinning behavior, with a resting viscosity of up to 200 Pa.s at a protein concentration of 20 mg/ml (Supplementary Fig. 9). This finding suggests that ALP could be well-suited for application via a spray-coating method, as the high shear rate at the spray nozzle could decrease the fluid’s viscosity, facilitating the formation of a thin and uniform coating on the fruit surface. Following the preparation of a uniform and compact ALP film on a polytetrafluoroethylene board with a thickness of 20 μm, it becomes evident that the ALP film possesses exceptional flexibility, capable of enduring repeated bending and folding without fracturing, showing the breaking strength and strain as 2.82 MPa and 39%, respectively (Fig. 3a, b). Such values are comparable to those of most edible films and the synthetic plastic films used in fruit packaging, such as pectin, starch, protein isolate, chitosan, carnauba wax, low-density polyethylene, and ethylene-vinyl alcohol copolymer7,25,26, indicating a potential for the ALP coating to resist premature failure or cracking during handling and storage. Furthermore, the ALP film exhibits high transmittance, being similar to the native protein adsorption layer (Fig. 3c), thereby preserving the natural appearance of the fruit.

Fig. 3: Characterization of the ALP coating to show multifunctional fresh-keeping features.
figure 3

a SEM images of ALP coating. Scale bar: 5 µm. b Mechanical properties of the ALP coating. c UV/vis transmission spectrum of the ALP coating, with the inset showing the ALP coating prepared on winter jujube. d The contact angle of a sessile water droplet on the ALP film as a function of time. e Water vapor transmission rate (WVTR) and Oxygen permeability (OP) of the ALP coating compared to common biopolymers used as a coating. f Respiration intensity of bare and ALP-coated cherry tomatoes at different storage times. g DPPH and ABTS radical scavenging activity of the ALP film. h Appearance of the ALP-coated and bare fresh-cut apple before and after 5 h of storage. i Antibacterial activity of lysozyme, cysteine, PTL, SA, CNC, and ALP on E. coli and S. aureus. j Weight loss and (k) stiffness of bare and lysozyme, cysteine, PTL, SA, CNC, lysozyme/SA, PTL/CNC, PTL/SA, and ALP-coated fresh-cut apples and winter jujubes at the end of storage. Different letters within each color indicate significant difference (p < 0.05). All data in Figs. 2f, g, 2i–k are mean ± S.D. n  =  3 independent samples per group. Statistical significance was determined by a two-tailed Student’s t test. The experiments in Fig. 2a were repeated independently at least three times with similar results. Source data are provided as a Source Data file.

Full size image

To gain further insights into the fresh-keeping features of ALP coating, we fabricated a self-supporting ALP film with about 28 µm thickness on a Teflon sheet. The contact angle of a water droplet on the ALP film, measured immediately after wetting, was approximately 75° (Fig. 3d), closely resembling the contact angle of 76° observed on chitosan films27. Notably, the contact angle decreased significantly from 71° to 60° over time, a phenomenon consistent with the time-dependent behavior observed on various polymer surfaces, likely attributable to surface remodeling induced by water absorption into the film27. Nonetheless, this coating material still exhibits greater hydrophobicity compared to some other polymers commonly employed for packaging, such as PET (52°), pullulan (30°), gelatin (65°), and pectin DE72 (54°)28. This heightened hydrophobicity may account for the low water permeability of the film, as fewer intermolecular interactions occur between water molecules and the hydrophobic surface, resulting in weaker capillary penetration and reduced water diffusion through the coating27. As expected, the water vapor transmission rate of the 57 µm-thick ALP film is then tested as approximately 4 g mm m−2 d−1, significantly lower than those of commonly used biopolymer packaging layers, such as polylactic acid (160 g mm m−2 d−1), SA (115 g mm m−2 d−1), pectin (75 g mm m−2 d−1), starch (78 g mm m−2 d−1), carboxymethyl cellulose (CMC) (65 g mm m−2 d−1), protein (180 g mm m−2 d−1), and gelatin-based composites29. This result thus supported the superior effectiveness of the ALP film in ensuring moisture retention within the fruit (Fig. 3e). In addition, the ALP film exhibits low oxygen permeability (OP) at 22 cm3 µm m−2 day−1 kPa−1, a value lower than that of other packaging materials (9–158 cm3 µm m−2 day−1 kPa−1). This result was ascribed to the incorporation of CNC, which resulted in a superiority in maintaining optimal oxygen barrier properties (Fig. 3e)30. This ensures the effectiveness of the film in delaying fruit ripening, as the lower oxygen levels in the microenvironment between the coating and the fruit reduce cellular respiration and slow down the ripening process. Benefiting from the above traits, it was then found that the ALP coating effectively reduced the respiration rate of cherry tomatoes (because of its minimal changes in respiration rate), delaying the arrival of peak respiration rate by 3 days, thereby extending their shelf-life (Fig. 3f).

Post-harvest fruits are easily oxidized when exposed to air, which causes nutrient loss, reduces skin resistance, and increases the susceptibility to microorganism invasion, especially in the case of fresh-cut fruits. As shown in Fig. 3g, the ALP coating exhibits excellent scavenging activity against both 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2′-casino-bis (3-ethylbenzothiazoline-6)-sulfonic acid (ABTS) radicals with the clearance ratio of 71% and nearly 100%, respectively (Supplementary Fig. 10). This capability is attributed to the presence of cysteine residues with the properties of antioxidation and polyphenol oxidase inhibition, along with the capacity of CNC within the ALP coating to capture and transfer electrons31. Upon application of the ALP coating to fresh-cut apples, it is evident that uncoated fresh-cut apples undergo significant browning, whereas those coated with the ALP maintain their original color even after 5 h of storage, highlighting the excellent antioxidative activity of the ALP coating (Fig. 3h). Besides, the ALP coating exhibited favorable antibacterial ability against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus), which can be assigned to the stronger and broader-spectrum antimicrobial properties of PTL24,32 (Fig. 3i and Supplementary Fig. 11). Based on the above advantages, the ALP coating thereby exhibits superior preservation effectiveness, as demonstrated by effectively preventing the decay and browning of winter jujube and fresh-cut apples (Supplementary Figs. 12 and 13). During storage, winter jujube was selected for the multi-component comparison experiment due to its longer shelf life. The fruits treated with the ALP coating exhibited less weight loss and maintained their stiffness, reflecting that the combined effect of the coating components effectively slowed down the weight loss and fruit softening (Fig. 3j, k and Supplementary Fig. 14).

Universal fruit’s fresh-keeping traits of the ALP coating

The preservation effect of the ALP coating on various types of perishable fruits was then validated, including non-climacteric and climacteric fruits. The results, as shown in Fig. 4a, revealed that bare strawberries exhibited noticeable signs of decay after 4 days of storage, while the ALP-coated strawberries remained intact at the same time. Subsequently, the coated strawberries showed no apparent decay or microbial damage until day 10, whereas all uncoated strawberries displayed severe fungal growth by that point (Fig. 4a and Supplementary Fig. 15). Similar to the preservation results observed for strawberries, the ALP coating extended the shelf-life of loquats from 4 days to 16 days, winter jujubes from 12 days to 21 days, and kumquats from 15 days to an impressive 30 days (Fig. 4a and Supplementary Figs. 16–18). Furthermore, the ALP coating was successfully applied to climacteric fruits, which featured high perishability and a brief shelf-life due to post-harvest physical and biological deterioration27. These fruits typically undergo a marked reduction in quality after reaching their peak respiration. Remarkably, the shelf-life of cherry tomatoes was prolonged from 6 days to an impressive 16 days, mangoes from 2 days to 8 days, nectarines from 2 days to 8 days, bananas from 2 days to 8 days, kiwis from 2 days to 8 days, ficus caricas from 1 day to 3 days, wolfberries from 1 day to 5 days (Fig. 4b and Supplementary Figs. 19–25). Furthermore, ALP coating demonstrates significant preservation efficacy under elevated temperatures. At 37 °C, the shelf-life of mangoes was extended by 3 days, and that of strawberries by 4 days. At 42 °C, the shelf-life of mangoes was extended by 2 days, and that of strawberries by 3 days (Supplementary Figs. S26, 27). These findings underscore the universal fresh-keeping capability of the ALP coating for fruits, doubling or even quintupling the typical shelf-life of highly perishable fruits (Fig. 4c).

Fig. 4: Effect of the ALP coating on ripening and sensory evaluation of non-climacteric and climacteric fruits.
figure 4

a Photographs of bare and ALP-coated (spray coating) strawberries, loquats, winter jujube, and kumquats after different storage times. b Photographs of bare and ALP-coated (spray coating) cherry tomatoes, mangoes, nectarines, and bananas after storage. c Comparison of shelf-life between ALP-coated and uncoated fruits. d LDA discriminant analysis of e-nose data for bare and coated strawberries at the end of storage. Source data are provided as a Source Data file.

Full size image

The good preservation effect of ALP can be attributed to its excellent film-forming ability, adhesion robustness, antioxidant properties, and the moisture barrier it provides. Furthermore, its antibacterial activity stands out as the most crucial factor in its preservative efficacy. To gain a more comprehensive understanding of the microorganisms associated with fruit spoilage across various fruit types and geographical regions, a systematic identification of the rot-causing strains in various fruits was conducted. The results showed that the dominant bacterial genera associated with fruit decay include pantoea in strawberry and nectarine, klebsiella in mango, dickeya in banana, gluconobacter in loquat, acetobacter in winter jujube, leuconostoc in loquat, and others. The prevalent fungal genera include fusarium in banana and kumquat, alternaria in strawberry, trichothecium in winter jujube, meyerozyma in loquat, mucor in cherry tomato and others (Supplementary Fig. 28). Given the broad-spectrum antibacterial properties of native lysozyme, representative bacteria responsible for fruit rotting were selected to further evaluate the antibacterial activity of ALP. The results showed that the PTL exhibited over 90% inhibition against mucor, fusarium, meyerozyma, gluconobacter and pantoea, while the PTL-containing ALP also demonstrated over 90% inhibition rate against meyerozyma, gluconobacter and pantoea. (Supplementary Fig. 29). In addition to its antimicrobial activity via lysozyme-mediated hydrolysis of peptidoglycan in the bacterial cell wall, the arginine and lysine residues on the surface of PTL used in the ALP coating increase the positive charge density (3.7 ± 0.2 mV), thereby enhancing the electrostatic interaction with the negatively charged bacterial membrane, leading to membrane disruption (Supplementary Fig. 30). Moreover, the phase transition of lysozyme exposes hydrophobic residues (Supplementary Fig. 30), which may further promote the destruction of the cell wall. Therefore, the combined effect of native lysozyme’s inherent antibacterial activity, the increased positive charge density in the PTL, and the exposed hydrophobic residues may contribute to its powerful antibacterial mechanism24,32.

Besides the appearance, the organoleptic properties of fruits were further assessed using an electronic nose (e-nose) and tongue (e-tongue) to evaluate the internal quality of the fruits. Taking the strawberries and cherry tomatoes as model perishable fruits, the radar chart clearly illustrated that the ALP-coated fruits maintained organoleptic qualities similar to fresh produce. In contrast, the uncoated samples showed significant changes in both aroma and flavor (Supplementary Figs. 31–34). Discriminant analysis utilizing linear discriminant analysis (LDA) revealed a noticeable difference in aroma and taste between the untreated group and the initially fresh fruits during storage (Fig. 4d and Supplementary Figs. 35–38). In contrast, fruits treated with the ALP coating retained aroma and taste characteristics comparable to fresh fruits throughout the entire storage period. Time-resolved LDA modeling on e-nose and e-tongue data further showed that at the end of storage, the aroma and taste from the ALP-treated fruits were very approaching to those from the uncoated fresh fruits before spoilage (Supplementary Figs. 39–41). These results suggest that the ALP coating not only prolongs the shelf-life of perishable fruits but also effectively maintains their aroma, nutritional value, and texture.

Furthermore, the changes of quality factors in the stored fruits, including weight loss ratio, stiffness, vitamin C (Vc) content, titratable acidity (TA), total soluble solids (TSS) content, and ultimately the statistical edible ratio, were then monitored to quantitatively validate the preservation effect of the ALP coating. Analysis of variance (one-way ANOVA) confirmed the homogeneity of the initial fruit samples, demonstrating no differences in their initial state (Supplementary Table 3). Specifically, across various fruits such as strawberries, loquats, winter jujube, kumquats, cherry tomatoes, mangoes, nectarines, and bananas, the ALP coating reduced weight loss by 33–69% by the end of the storage period in comparison to the uncoated fruits (Fig. 5a, b and Supplementary Fig. 42). This reduction is attributed to the ALP coating’s ability to inhibit water vapor transmission. In addition to weight loss, the stiffness loss of ALP-coated fruits was also reduced by 9%–91% compared to uncoated fruits (Fig. 5a, c, and Supplementary Fig. 43). Moreover, the loss of Vc in the ALP-coated non-climacteric fruits (e.g., strawberries, loquats, kumquats, and winter jujubes) was reduced by 42%–92% compared to the bare fruits (Fig. 5a, d and Supplementary Fig. 44). This trend was also consistent in the ALP-coated climacteric fruits (e.g., cherry tomatoes, mangoes, bananas, and nectarines), with a 14% ~ 39% reduction in Vc loss compared to the uncoated fruits (Supplementary Fig. 44). As an indication of the flavor quality of fruits, the ALP coating was then found to effectively reduce TA loss by 21–65% compared to uncoated fruits (Fig. 5a, e, and Supplementary Fig. 45). Similarly, the reduction in TSS was decelerated in all ALP-coated fruits compared to uncoated fruits, reducing loss by 2–89% (Fig. 5a, f, and Supplementary Fig. 46). Climacteric fruits, such as cherry tomatoes, exhibit an initial increase followed by a decrease in Vitamin C, TA, and TSS content. This pattern is attributed to their continued ripening post-harvest, characterized by a respiratory climacteric and increased ethylene production. This respiratory burst ultimately leads to accelerated degradation of these compounds. Conversely, non-climacteric fruits, such as strawberries, do not undergo respiratory climacteric, and their ripening process significantly slows after harvest. More intriguingly, the application of ALP coating on fruits delayed the onset of the respiratory peak, leading to reduced erosion of Vc, TA, and TSS (Supplementary Fig. 47). Based on the above collective evaluations, the edible ratio of the fruits further confirmed a significant increase of 30-90% in freshness index compared to uncoated fruits (Fig. 5g and Supplementary Fig. 48). In addition, the thickness and root mean square roughness (RMS) of the coatings obtained by the spraying and dipping did not differ significantly (Supplementary Fig. 49), and the shelf-life and nutrient retention of these two ALP coatings showed no significant difference, demonstrating that both methods are suitable for subsequent practical production (Fig. 5b–g and Supplementary Figs. 15–22).

Fig. 5: Effect of the ALP coating on ripening and quality of fruits.
figure 5

a ALP-coated fruits exhibit a significant reduction in the rate of weight loss and demonstrate enhanced preservation of stiffness, vitamin C (Vc), titratable acid (TA), and total soluble solids (TSS) relative to their uncoated counterparts. Data are mean ± S.D. n  =  3 independent samples per group. b Weight loss, (c) stiffness, (d) Vc content, (e) TA content, (f) TSS content, and (g) edible rate of bare and coated strawberries at different storage times at 23 °C and 50 % humidity. All data in Fig. 4b–g are mean ± S.D. n  =  3 independent samples per group. Statistical significance was determined by one-way ANOVA with Tukey’s multiple comparison test. Details (p-value) of statistical comparisons between groups are provided in the Supplementary appendix. Source data are provided as a Source Data file.

Full size image

The suitability to employ the ALP coating for the preservation of fresh-cut fruits

Encouraged by the aforementioned results on post-harvest fruits, the effectiveness of the ALP coating on freshly cut fruits was further examined. This is particularly important as cut and peeled fruits tend to experience quicker quality deterioration than unpeeled fruits, often manifesting as moisture loss, softening, enzymatic browning, microbial contamination, and aroma loss. Notably, as illustrated in Fig. 6a, the ALP-coated fresh-cut apples exhibited a lower browning index (BI) compared to their untreated counterparts. Evaluation of key parameters such as weight loss, stiffness, the content of Vc, TA, and TSS for the ALP-coated fresh-cut apples at different storage intervals revealed significant improvements over the untreated fresh-cut apples (Supplementary Fig. 50). Specifically, the weight loss of the ALP-coated apples was reduced by 88% compared to untreated apples, with nutrient retention ranging from 57% to 86%, as opposed to only 8% to 39% in the untreated group. The e-tongue and e-nose data further indicated that, by the end of the storage period, the scent and taste of the ALP-treated fresh-cut apples closely mirrored those of fresh apples (Fig. 6b-d and Supplementary Figs. 51–55). In addition, the ALP-coated fresh-cut apples exhibited bacterial presence on day 8 and fungal growth on day 12, whereas untreated fresh-cut apples showed signs of bacterial presence on day 2 and fungal growth on day 4, highlighting the antimicrobial properties of the ALP (Supplementary Figs. 56, 57). Furthermore, our preservation experiment involving a variety of fresh-cut fruits—such as apples, cherry tomatoes, strawberries, bananas, dragon fruit, mangoes, grapes, and blueberries—revealed significant discrepancies in shelf-life. By day 4, the untreated fruit assortment began to display browning and decay, whereas the ALP-coated assortment maintained a high level of freshness up to day 10, indicating a 4-fold extension in shelf-life (Fig. 6e and Supplementary Fig. 58). Besides, the ALP treatment maintained considerable preservation efficacy even under extreme high-temperature conditions (42 °C), extending the 2-fold shelf-life of fresh-cut apples (Supplementary Figs. 59, 60). In conclusion, ALP coating effectively preserves the flavor and nutritional quality of fresh-cut fruits, highlighting its considerable potential in fruit preservation technology.

Fig. 6: The application of the ALP coating on the ripening and shelf-life of fresh-cut fruits.
figure 6

a The impact of the ALP treatment on the appearance quality of fresh-cut apples was assessed at 4 °C and 50% humidity after 0, 6, and 10 days of storage. b Radar chart of e-nose data of bare and coated fresh-cut apples obtained at the end of storage. c LDA discriminant analysis of e-nose data for bare and coated fresh-cut apples at the end of storage. d Predictive results of e-nose data at the end of storage for coated fresh-cut apples in the LDA model. e The influence of the ALP treatment on the visual quality of fresh-cut fruit assortments was evaluated over a storage duration ranging from 2 to 10 days. Source data are provided as a Source Data file.

Full size image

Washable, edible, and sustainable properties of the ALP coatings

While the ALP coating consists of amyloid protein and natural products deemed safe for consumption33,34,35, we conducted a Congo Red staining test to assess the washability of the coating when not intended for ingestion. Following rinsing of the coated surface with water, fluorescence microscopy revealed the absence of Congo Red, affirming the coating’s ease of cleaning (Fig. 7a), which is largely ascribed to the non-covalent interactions of the ALP coating with the fruit surface. The contact angle measurements taken on tomato and banana surfaces before and after cleaning further supported the removal of the ALP coating from the fruit peel (Fig. 7b). The ALP coating was then spray-coated onto rat food, thoroughly dried, and administered to two groups of SD rats under the GB 15193.22-2014 standard, with observations spanning 28 days. Throughout the observation period, we observed no significant differences in food and water consumption, no mortality in the ALP-coated group, and no notable disparities in body weight changes between the two groups (Supplementary Fig. 61). Organ and blood samples were collected after the 28-day experimental phase for pathological examination, routine blood tests, as well as liver and kidney function assessments. As shown in Fig. 7c and Supplementary Fig. 61, no distinct pathological alterations were observed in the pathological sections, and there were no discernible differences between the ALP-treated group and the control group. Furthermore, the results of routine blood tests and liver and kidney function tests fell within normal ranges (Fig. 7d–f). Besides, the hemolysis test indicated that the hemolysis ratio of ALP was 2.46%, well below the ASTM standard (ASTM F756-2008, < 5.00%), demonstrating the excellent biocompatibility of the ALP coating (Supplementary Fig. 62). Collectively, these outcomes demonstrate the favorable biosafety profile of the ALP coating and warrant its safety for consumption.

Fig. 7: The biosafety assessment of the ALP coating.
figure 7

a Fluorescent images showing the ALP coating before and after cleaning with Congo red staining. b Water contact angle measurements on ALP-coated cherry tomatoes and bananas before and after cleaning. c Pathological examination results of heart, liver, spleen, lung, kidney, and intestine tissues. The control group was fed bare rat food, while the coated group was fed ALP-coated rat food. Results of (d) blood routine (white blood cell (WBC), neutrophil (Neu), lymphocyte (Lym), (e) liver function (alanine aminotransferase (ALT), aspartate transaminase (AST)), and (f) kidney function tests in both control and coated groups are presented (creatinine (CREA), blood urea nitrogen (BUN)). n  =  6 animals per group. Data are presented as a box and whisker plot (median, box:first and third quartiles, and whisker: minimum and maximum). All data in Fig. 6d–f are mean ± S.D. Statistical significance was determined by a two-tailed Student’s t test. The experiments in (a, c) were repeated independently at least three times with similar results. Source data are provided as a Source Data file.

Full size image

A comprehensive life cycle assessment (LCA) was then conducted to evaluate the environmental impact and sustainability of the ALP coating as a method of fruit preservation. This analysis compared the carbon footprint resulting from processing 1 kg of ALP-coated fruits at ambient temperature (23 °C) against 1 kg of uncoated fruits stored in a refrigerator at 4 °C. The LCA revealed that 1 kg of refrigerated cherry tomatoes at 4 °C began to spoil on the fourth day, generating 0.055 kg of CO2 emissions (Fig. 8a, b and Supplementary Table 4). In contrast, 1 kg of the ALP-coated cherry tomatoes stored at 23 °C demonstrated a significantly extended shelf-life of 10 days, only producing 10% CO2 emissions of those from the refrigerated sample over the same period (Fig. 8b, Supplementary Fig. 63, and Supplementary Table 4). The assessment also encompassed non-climacteric fruits like strawberries, where the ALP-coated samples demonstrated preservation efficacy on par with refrigerated counterparts (Supplementary Fig. 64). In such a case, the difference in weight loss, hardness, Vc, TA, and TSS loss between the ALP-coated and refrigerated fruits was only about 1% (Fig. 8c, d and Supplementary Figs. 65, 66). The above results thus highlighted the potential of ALP coating as a viable alternative to refrigeration for fruit preservation, offering a substantial reduction in carbon emissions.

Fig. 8: The sustainable assessment of ALP.
figure 8

a Images showcasing cherry tomatoes stored in the fridge at 4 °C and coated at room temperature (23 °C and 50% humidity) on days 0, 4, and 8. b Life Cycle Assessment (LCA) of the ALP coating for preserving cherry tomatoes at room temperature (23 °C and 50% humidity) compared to refrigeration at 4 °C. The (c) weight loss and (d) Vc content of cherry tomatoes stored in the fridge at 4 °C and coated at room temperature (23 °C and 50% humidity) at different storage durations. Data are mean ± S.D. n  =  3 independent samples per group. Statistical significance was determined by one-way ANOVA with Tukey’s multiple comparison test. Source data are provided as a Source Data file.

Full size image

In addition to refrigerating fruits for storage, our comparative analysis evaluated the effectiveness, versatility, and safety of the ALP coating in comparison to other traditional preservation methods, e.g., fruit waxing as the commonly used commercial technique for fruit preservation (Table 1). In addition, various transparent films, such as those composed of pea starch, chitosan, konjac glucomannan, and pullulan, combined with antimicrobials and antioxidants, are also included for comparison, although their broad applicability and biosafety are still in question. It is worth noting that while traditional methods are typically limited to specific fruits, the ALP method has demonstrated universal applicability across a wide range of fruit types, including both climacteric and non-climacteric varieties. Notably, by taking strawberries as a specific example, the ALP coating has significantly extended its shelf-life by up to five times, outperforming other coatings documented in the current literature (Table 1). The superior performance of the amyloid-like protein (ALP) coating stems from its enhanced spreadability, film formation, adhesion robustness and uniformity on fruit surfaces, which result in improved coating stability and prolonged preservation. Conventional coatings often exhibit uneven adhesion and poor stability, particularly on hydrophobic peels. For instance, coatings incorporating clove essential oil or nisin with sodium alginate/cellulose nanocrystals (SA/CNC) demonstrate suboptimal film formation on hydrophobic tomato skin (Supplementary Figs. 67, 68) and lower adhesion strength compared to ALP (Supplementary Fig. 69). Consequently, these coatings only achieve a 2-fold shelf-life extension, whereas ALP achieves a 5-fold extension (Supplementary Fig. 70). In addition, preservation experiments were conducted by replacing SA in the ALP coating with starch, gelatin, and chitosan, respectively. The results showed that starch, gelatin, and chitosan only extended the shelf life by 2 times, while the preservation effect of ALP (SA) was significantly better than the others (Supplementary Fig. 71), further demonstrating the excellent preservation ability of ALP (doped with SA). It is then crucial to note that although waxing is a widely adopted method, it has elicited concerns regarding its effects on long-term consumption and health implications, particularly due to its potential to induce anaerobic respiration and restrict O2 and CO2 exchange36. The biocompatible, edible, and easily washable nature of the ALP coating provides a practical and safer alternative for food preservation. Besides, our analysis suggests that the ALP preservation coating is cost-efficient, requiring only $0.09 per kilogram of fruit. This cost-effectiveness, combined with its superior fresh-keeping properties and broad applicability, underscores the great potential of the ALP strategy as a significant advancement in fruit preservation methods.

Table 1 Application range and shelf-life extension of various preservation coatings
Full size table

Perishable fruits contribute significantly to annual food waste due to the limited effectiveness of their natural cuticles in extending shelf life and storage potential. To address this issue, eco-friendly amyloid-like protein (ALP) coatings have been developed in this work for fruit preservation. These coatings involve the formation of a dense, cling phase-transitioned protein/polysaccharide composite film on the fruit’s epicuticular cuticles. The application of these coatings through immersion and spraying techniques is universal across a dozen fruit types. The ALP strategy applies to 17 different families of perishable fruits and fresh-cut fruits, encompassing both non-climacteric and climacteric species such as strawberries, loquats, kumquats, jujubes, mangoes, bananas, cherry tomatoes, nectarines, kiwis, ficus carica, and wolfberries. The ALP coating significantly extends the shelf life of fruits by inhibiting microbial growth and reducing moisture loss, thereby delaying decay. Notably, the ALP coating has been shown to increase the shelf-life of these perishable items by 2 to 5 times while preserving 60–98% of nutrients until the end of the storage period. The edibility, ease of removal, cost-effectiveness, and suitability of the ALP coating for preserving fresh-cut fruits have been confirmed through its application on fruit platters. It offers greater generalizability, improved freshness retention, and enhanced biocompatibility compared to traditional preservation methods. In addition, the ALP coating can replace the cold chain storage of fruits, leading to a 90% reduction in carbon emissions while providing a 2.5-fold longer shelf-life compared to refrigerated storage. In conclusion, the ALP system emerges as a promising and sustainable approach to extending the shelf-life of perishable fruits, with the potential to significantly reduce food waste within the industry and in life.

Methods

Materials

Lysozyme (extracted from egg white) was purchased from Sigma-Aldrich. L-cysteine was obtained from Aladdin. Sodium alginate (SA) was purchased from Aladdin. Glycerol (95.5 ≥ purity) was purchased from Sigma-Aldrich. Phosphate-buffered saline (PBS), Mueller-Hinton broth (MHB), and Mueller-Hinton agar (MHA) were purchased from Solarbio. Thioflavin T (ThT) and 1-anilino-8-naphthalenesulfonic acid (ANS) were purchased from Sigma-Aldrich. Staphylococcus aureus (S. aureus) (ATCC6538), Escherichia coli (E. coli) (ATCC25922), and L929 cells were obtained from the American Type Culture Collection (USA). SYTO 9 green fluorescent nucleic acid stain was purchased from Thermo Fisher. Winter jujube, strawberries, and tomatoes were sourced from Shaanxi, China; loquats from Sichuan, China; kumquats and mangoes from Guangxi, China; and bananas from Hainan, China.

Measurements and characterizations

Scanning electron microscopy (SEM) was conducted on an FEI Quanta 200 electron microscope (SU8020, Hitachi) with a primary electron energy of 5 kV. Far-UV circular dichroism (CD) spectrum was recorded by using a Chirascan spectrophotometer (Applied Photophysics Ltd, England). The water contact angle (WCA) was performed by an interface/tensiometer (DCAT 21 and OCA 20, Dataphysics). The Fourier transform infrared (FTIR) spectra were recorded on a Tensor 27 (Bruck) spectrometer over the range of 4000–600 cm−1 to analyze the functional groups of the materials. The transmittance of the amyloid-like protein coating adhered to a quartz plate was collected by a U-3900/3900H (Hitachi). The X-ray photoelectron spectroscopy (XPS) spectra were obtained using an X-ray photoelectron spectrometer (AXIS ULTRA, Kratos Analytical Ltd.). The binding energies were calibrated by setting the C1s peak at 284.6 eV. The fluorescence spectra were collected by an F-7000 fluorescence spectrophotometer (Hitachi). Biological samples were fixed with 4% paraformaldehyde and dehydrated before observation.

Preparation of the amyloid-like protein solution

The phase transition solution, typically used for this purpose, consists of lysozyme, sodium alginate, and cellulose nanocrystals. To prepare the solution, an aqueous solution of lysozyme (10 mg/mL) is mixed with sodium alginate (10 mg/mL, containing 0.3% glycerol) and cellulose nanocrystal (0.1 wt%). Then, a cysteine solution (10 mg/mL, pH 8) is added, followed by stirring for five minutes.

Preparation of amyloid-like protein coating on fruits

The nanocomposite solution can be applied to fruits using either dip-coating or spray-coating methods. Various fruits, including winter jujube, strawberry, kumquat, loquat, nectarine, cherry tomato, banana, mango, kiwi, ficus carica, wolfberry, and fresh-cut fruit, were fully immersed in the solution or sprayed with it. After a 2-minute soaking period, the fruits were coated with a layer and then air-dried at room temperature.

Mechanical properties

The tensile strength and elongation at the break of the composite films were determined using a Tensile Testing Machine (Sansi Technology Co., Ltd., Shenzhen, China) with initial grips separation set at 20 mm and a probe speed of 1 mm/s. Samples, cut into 50 mm × 10 mm strips, were tested in a natural environment with a temperature of approximately 23 °C and humidity around 50%. The measurements were conducted three times, and the average value was recorded.

180° peel test

The sample was affixed to a stainless-steel plate, and the ALP-coated side was secured with double-sided adhesive tape. A 10 kg weight was then applied to the sample for 24 h. After removing the weight, a 180° peel test was conducted using a tension machine (Sansi Technology Co., Ltd., Shenzhen, China).

Weight loss

The weight loss of the fruits was determined by calculating the difference between the initial weight and the weight measured with a precision of 0.0001 g at daily intervals. The weight loss rate was obtained using Eq. (1).

$$W=\frac{{m}_{0}-{m}_{n}}{{m}_{0}}\times 100\%$$
(1)

where \({m}_{0}\) (g) is the initial mass of fruits; \({m}_{n}\) (g) is the mass of fruits after n days.

Stiffness: The firmness of the fruits was assessed using a hand-held fruit firmness tester (GY-4, Guangzhou, China). Following the guidelines specified in Chinese NY/T 2009-2011, a probe with a diameter of 8 mm was used to measure the firmness at a designated point in the central region of each fruit.

Edible ratio

The edible ratio of the fruits was determined by calculating the difference between the initial fresh fruit quantity and the fresh fruit quantity counted each day. The edible ratio is derived from the equation:

$${{{\rm{Edible}}}}\; {{{\rm{ratio}}}}=\frac{{N}_{n}}{{N}_{0}}\times 100\%$$
(2)

where \({N}_{0}\) is the initial number of fresh fruits; \({N}_{n}\) is the number of fresh fruits after day n.

Determination of vitamin C

The ascorbic acid content in fruits was measured following the GB 5009.86-2016 standard using a titration technique. Initially, 20.00 g of fruit homogenate was placed in a 100 mL volumetric flask, and the volume was made up to 100 mL with a 20 g/L oxalic acid solution. Next, 8 g of kaolin was added to the solution for decolorization, followed by filtration. A 10.0 mL sample of the filtrate was titrated with a 0.1 mg/mL 2,6-dichlorophenol solution until a pink color appeared. The ascorbic acid content was then determined using Eq. (3):

$$\,X=\frac{(V-{V}_{0})\times T\times A}{m}\times 100$$
(3)

where \(X\) (mg/100 g) represents the content of L (+)-ascorbic acid in the sample; \(V\) (mL) represents the volume of 2, 6-dichlorophenol solution consumed by titrating the sample; \({V}_{0}\) (mL) represents the volume of 2, 6-dichlorophenol solution consumed by titrating blank; \(T\) (mg/mL) represents the titration of 2, 6-dichlorophenol solution; \(A\) is the dilution; \(m\) (g) represents the sample quality.

Determination of titratable acid

The titratable acid content of fruits was determined using the titration method according to GB/T 12456-2008. After grinding, 20.00 g of fruit homogenate was transferred to a 100 mL volumetric flask, and the volume was adjusted to the mark with water. To 10.0 mL of the filtrate, 0.2 mL of phenolphthalein indicator was added. The solution was then titrated with a 0.1 mol/L NaOH solution until a pink color appeared. The titratable acid content (TA) was calculated using Eq. (4):

$${TA}=\frac{C\times ({V}_{1}-{V}_{2})\times K\times F}{m}\times 100\%$$
(4)

where \({TA}\) (%) denotes the titratable acid content of strawberries; c (mol/L) represents the concentration of NaOH; \({V}_{1}\)(mL) represents the volume of NaOH solution consumed by the test solution; \({V}_{2}\)(mL) represents the volume of NaOH solution consumed of blank; \(K\) denotes the conversion coefficient of acid, 0.067; \(F\) denotes the dilution ratio; \(m\) (g) represents the mass of the sample.

Determination of total soluble solids

The soluble solids content of fruits was determined using a refractometer, following the GB/T 12295-90 standard. After mashing the fruits, the homogenate was filtered through gauze, and the resulting juice was measured using an Abbe refractometer (WAY-2WAJ, Shanghai, China).

Browning index analysis (BI)

The L*, a*, and b* values of the simulated system were measured using a WSD-3C automatic whiteness colorimeter. A positive L* value indicated the brightness of the object, while positive values of a* and b* represented the redness and yellowness of the samples, respectively. Each sample was measured in triplicate. The Browning Index (BI) of the simulated system was calculated using the following formulas:

$${BI}=\frac{100\times (x-0.31)}{0.172}$$
(5)

where,

$$x=\frac{{a}^{*}+0.175{L}^{*}}{5.645{L}^{*}+{a}^{*}-3.012{b}^{*}}$$
(6)

Calculation of shelf-life

The shelf-life of the fruit is determined based on the last day when it remains unspoiled. The formula for calculating the extended shelf-life multiplier

$${{{\rm{shelf}}}}\; {{{\rm{life}}}}\,\,{{{\rm{multiplier}}}}=\frac{{S}_{{ALP}}}{{S}_{{bare}}}$$
(7)

Where \({S}_{{ALP}}\) is the shelf-life of ALP coated fruit and \({S}_{{bare}}\) is the shelf-life of bare fruit.

Molecular dynamics simulation

Molecular dynamics (MD) simulations were conducted with the Gromacs 2023.2 package37 with the Amber ff14SB force field38. To investigate the structure of cuticular wax, we constructed a hydrophobic complex mixture composed of long-chain fatty acids, corresponding derivatives, and triterpenoids. The composition of the mixture included 9(10),16-dihydroxyhexadecanoic acid, 18-hydroxy-9,10-epoxyoctadecanoic acid, hentriacontane, hexacosanoic acid, p-coumaric acid, and ursolic acid. The cuticular wax structure was built by the Packmol program39 by inserting the above molecules with an amount of 200, 200, 300, 200, 200, and 100 (controlling the component as the natural cuticular wax) into a 140 × 114 × 51.4 Å rectangle box. The cuticular wax structure was first equilibrated at 363.15 K for 40 ns. Then the lysozyme at native state (PDB entry: 1LYZ) and fibril (PDB entry: 8QV8) states were each positioned 5 Å and 2 Å above the equilibrated cuticular wax complex. The topology of each lysozyme was generated with all disulfide bonds cleaved. Then the systems underwent 2000 energy minimization steps and a 200 ns product phase simulation. The MD simulations employed an NVT ensemble utilizing a time step of 2 fs, maintaining a constant temperature of 298.15 K, controlled by the velocity-rescale method40. The temperature coupling constant was set to 0.2 ps. Visualization of the simulation snapshots was performed using PyMOL (www.pymol.org) and VMD software41. The binding energy between the lysozyme and cuticular wax was analyzed by the s_mmpbsa program42, excluding the PBSA calculation (in NVT ensemble).

Contact angle measurement

The static contact angle (CA) of a liquid droplet was measured with a contact angle meter. The volume of the liquid droplets was controlled to be approximately 2 μL, and each sample was tested at least three times.

Fourier transforms infrared (FT-IR) spectroscopy

FT-IR spectra of the prepared ALP and native protein adhesive were recorded on a spectrometer (Thermo Scientific Nicolet iS50, America). Samples were mixed with dried KBr and then pressed into pellets for FTIR analysis performed in the range from 4000 cm−1 to 500 cm−1 at a scanning rate of 4 cm−1.

Scanning electron microscopy (SEM)

ALP films were observed on an FEI Quanta 200 electron microscope (SU8020, Hitachi) with a primary electron energy of 5 kV. Before analysis, the samples were coated with gold.

Water vapor permeability (WVP)

The gravimetric method was employed to investigate the WVP of the film samples29. The films were cut into suitable sizes and fixed onto the mouth of glass cups (internal diameter of 1.1 cm and a depth of 6.3 cm), which contained about 30 g anhydrous calcium chloride (CaCl2) with ~0% RH. Thereafter, the cups were transferred into a desiccator which was saturated with NaCl (75% RH) solution and kept for 24 h at 25 °C. The changes in the weight of the cups were periodically recorded every 1 h during the first 8 h and finally after 24 h. The WVP was determined with Eq. (8):

$${WVP}=\frac{\triangle w\times X}{t\times A\times \triangle P}$$
(8)

where \(\triangle w\) is the weight change in the cup (g), \(X\) is the film thickness (m), t is the time (s), \(A\) is the effective area of the film (m2), and \(\triangle P\) is the water vapor pressure difference on each side of the film.

Oxygen permeability (OP)

The oxygen permeability of ALP composite films was measured by the deoxidizer absorption method43. Films were sealed on the top of test cups with an internal diameter of 25 mm and a depth of 40 mm, filled with a certain amount of deoxidizer (reduced iron powder). Then, the cups were placed in the desiccator with a temperature of 23 ± 2 °C and a relative humidity of 75 ± 5 % for 48 h. The OP value was determined by the following formula (9):

$${OP}=\frac{\triangle m}{t\times A}$$
(9)

where, OP is the oxygen transmission rate of composite films, \(\triangle m\) (g) is the weight change of the est cup before and after the test, t (h) is the test time and \(A\) (m2) is the test area. The measurements were repeated three times, and the average value was recorded.

Antioxidation properties

Two simulated food environments were used to evaluate the free radical scavenging effects of the composite film44. Briefly, 50 mg of the film was dissolved in ethanol solutions (10 mL) and stirred on a magnetic stirrer. After 3 h, a sample of the supernatant was centrifuged and used for the measurement. The results of the analysis were compared to those for the ALP film. The DPPH solution was mixed with the film extract solution and left in the dark for 1 h. The absorbances of the samples were measured at 517 nm, and the DPPH scavenging activity was calculated using Eq. (10):

$${DPPH\; radical\; scavenging\; activity}\,\,(\%)=\frac{{A}_{{control}}-{A}_{{sample}}}{{A}_{{control}}}\times 100$$
(10)

A mixture of 0.737 mL ABTS solution (7.4 mmol methanol solution) and 1.43 mL potassium persulfate solution (7.4 mmol aqueous solution) was allowed to react for 12 h in the dark. When the mixture solutions were diluted 20 times with distilled water, the absorbance of the mixture at 734 nm remained constant at 0.7. Then, the ABTS solution was mixed with the extracts from the film and left in the dark for 10 min. The absorbance of the samples was read at 734 nm, and the ABTS scavenging activity was calculated using Eq. (11):

$${ABTS\; radical\; scavenging\; activity}\,\,(\%)=\frac{{A}_{{control}}-{A}_{{sample}}}{{A}_{{control}}}\times 100$$
(11)

Bacterial inhibition ratio measurement

Antibacterial activity of the ALP was evaluated based on the colony counting method. Two bacteria were used in this study, including Staphylococcus aureus (S. aureus, ATCC 6538) and Escherichia coli (E. coli, ATCC 25922). At first, the bacteria were cultured aerobically in 50 mL MHB overnight at 37 °C under shaking at 70 rpm, so that the bacteria were in the logarithmic growth phase. 1 mL of the bacterial suspension was collected in a sterile centrifuge tube, centrifuged at 5000 rpm, and washed with PBS to remove the culture medium. This process was repeated three times. Finally, the test bacteria were re-suspended in PBS at a concentration of 105 CFU mL− 1. Then, adding 10 μL the above bacterial suspension was incubated in 1 mL Lysozyme, Cysteine, PTL, SA, CNC, ALP, and PBS for 8 h in a humid chamber at 37 °C. The bacterial suspensions were diluted and spread on MHA plates for the plate counting assay. After 24 h of culture growth at 37 °C, the total number of bacterial colonies (C) in each plate was recorded. Finally, the antibacterial activity is represented by the killing ratio, which is calculated according to the following Eq. (12):

$${Inhibition\; ratio}=\frac{{C}_{0}-C}{{C}_{0}}\times 100\%$$
(12)

where C0 is the bacterial colony of the control group, C is the bacterial colony of the experimental group. The measurement was repeated five times, and the average was regarded as the final result.

Hemolysis assay

All blood experiments were approved by the Biomedical Ethics Committee of Health Science Center of Xi’an Jiaotong University (No. XJTUAE2023-942). Fresh blood with EDTA anticoagulant was centrifuged at 3000 rpm for 5 min. The supernatant was then removed, and the lower layer of red blood cells was washed three times with PBS. The blood cells were then mixed with PBS in a 1:1 ratio to dilute the red blood cells. 2 mL of PBS, ALP, and 40 μL of blood cells were added to the experimental group; 2 mL of PBS and 40 μL of blood cells were added to the negative control group; and 2 mL of ultrapure water and 40 μL of blood cells were added to the positive control group. The hemolysis ratio of the samples can be calculated using the following formula:

$${{HR}}=\frac{A-B}{C-B}\times 100\%$$
(13)

HR is the hemolysis ratio, A, B, and C represent the absorbance values of the experimental group, the negative control group (there is no red blood cell rupture in normal saline), and the positive control group (the red blood cell ruptures in water, resulting in the release of a large amount of hemoglobin).

Animal experiment

All experiments were performed according to the guidelines of the Biomedical Ethics Committee of Health Science Center of Xi’an Jiaotong University (No. XJTUAE2023-942). All animals are 6-week-old male SD rats, weighing approximately 200 g and housed in an experimental animal room maintained on a 12:12 h, light:dark cycle. The temperature and humidity were maintained ~ 20 °C and between 45–55%, respectively. The animal toxicity assessment of ALP is performed according to the following steps. First, ALP coating is sprayed onto the rats’ food and thoroughly dried. The two groups of SD rats are then fed according to the GB 15193.22-2014 standard, with an observation period of 28 days. During the entire observation period, food and water are provided abundantly, and body weight is measured every 7 days. After 28 days, the rats are sacrificed, and major organs such as the stomach and intestines are collected and fixed in a 4% paraformaldehyde solution for tissue section staining analysis. In addition, blood samples are collected for routine blood tests and liver and kidney function assessments.

Life cycle assessment (LCA)

A complete life cycle assessment includes the process from the extraction of product raw materials to product processing, transportation, and degradation45. The preparation process of ALP involves lysozyme, cysteine, SA, and CNCs. Since it cannot be obtained from the source of materials, we have replaced some data with the relevant literature and existing data in the Ecoinvent (v3.8) database (https://www.globallcadataaccess.org/, https://ecoinvent.org/). The analysis follows the ISO standard 14040 of LCA and the ISO standard 17067 for the carbon footprint. In addition, SimaproTM (v9.4) was used to simulate the environmental impact of ALP. The data used in the simulation, in which ALP coating is prepared by spraying.

Statistical analysis

All data are mean ± S.D. Student’s t test (two-group comparison), one-way ANOVA, or two-way ANOVA (multiple-groups comparison) were performed to determine statistical significance between different groups. Statistical tests and exact P-values used for each data are presented in the figure and legend. Statistics were calculated using GraphPad Prism 9.

Reporting summary

Further information on research design is available in the Nature Portfolio Reporting Summary linked to this article.